A Lipolytic Lecithin:Cholesterol Acyltransferase Secreted by Toxoplasma Facilitates Parasite Replication and Egress

Abstract The protozoan parasite Toxoplasma gondii develops within a parasitophorous vacuole (PV) in mammalian cells, from where it scavenges cholesterol. When cholesterol is present in excess in its environment, the parasite expulses this lipid into the PV or esterifies it for storage in lipid bodies. Here, we characterized a unique T. gondii homologue of mammalian lecithin:cholesterol acyltransferase (LCAT), a key enzyme that produces cholesteryl esters via transfer of acyl groups from phospholipids to the 3-OH of free cholesterol, leading to the removal of excess cholesterol from tissues. TgLCAT contains a motif characteristic of serine lipases AHSLG and the catalytic triad consisting of serine, aspartate and histidine [SDH] of LCAT enzymes. TgLCAT is secreted by the parasite, but unlike other LCAT enzymes, is cleaved into two proteolytic fragments that share the residues of the catalytic triad and that need to be reassembled to reconstitute enzymatic activity. TgLCAT uses phosphatidylcholine as substrate to form lysophosphatidylcholine that has the potential to disrupt membranes. The released fatty acid is transferred to cholesterol, although with a lower transesterification activity than mammalian LCAT. TgLCAT is stored in a subpopulation of dense granule secretory organelles, and following secretion, it localizes to the PV and parasite plasma membrane. LCAT-null parasites have impaired growth in vitro, reduced virulence in animals and exhibit delay in egress from host cells. Parasites overexpressing LCAT show increased virulence and faster egress. These observations demonstrate that TgLCAT influences the outcome of an infection, presumably by facilitating replication and egress depending on the parasite developmental stage.